enhancer capture Hi-C of Mus musculus: embryonic day 11.5 neural tube
Briefly, frozen nuclei pellets (2-6 million) were thawed on ice, followed by adding SDS and Triton X-100 to remove non-crosslinked proteins and sequester SDS, and digested using DpnII (NBE) overnight at 37 . The ends of restriction fragments were labeled with biotinylated dCTP and ligated at room temperature for 4 hours. After de-crosslinking and precipitation, ligated products were sheared using a Covaris sonicator (duty cycle: 10%, intensity: 5, cycles per burst: 200, treatment time: 180s in total) to an average fragment size of 200bp. The ligated sheared 3C libraries (10-12 ug for each replicate) were pulled down using Streptavidin Dynabeads (Thermo Scientific) to get rid of unligated fragments, followed by end repair, adaptor ligation and amplification according to modified Agilent SureSelectXT protocol. A target-enrichment was performed using capture RNA probes according to Agilent SureSelect XT protocol with an input amount of 750 ng of Hi-C library per sample. After indexing and amplification, libraries were sequenced (paired-end 100 or 150 bp) on an Illumina NovaSeq 6000.